Plyler Jason, Jasheway Karl, Tuesuwan Bodin, Karr Jessica, Brennan Jarryd S, Kerwin Sean M, David Wendi M
Department of Chemistry and Biochemistry, Texas State University-San Marcos, San Marcos, TX 78666, USA.
Cell Biochem Biophys. 2009;53(1):43-52. doi: 10.1007/s12013-008-9038-z. Epub 2008 Dec 2.
The simian virus 40 (SV40) genome is a model system frequently employed for investigating eukaryotic replication. Large T-antigen (T-ag) is a viral protein responsible for unwinding the SV40 genome and recruiting necessary host factors prior to replication. In addition to duplex unwinding T-ag possesses G-quadruplex DNA helicase activity, the physiological consequence of which is unclear. However, formation of G-quadruplex DNA structures may be involved in genome maintenance and function, and helicase activity to resolve these structures may be necessary for efficient replication. We report the first real-time investigation of SV40 T-ag helicase activity using surface plasmon resonance (SPR). In the presence of ATP, T-ag was observed to bind to immobilized single-stranded DNA, forked duplex DNA, and the human telomeric foldover quadruplex DNA sequence. Inhibition of T-ag duplex helicase activity was observable in real-time and the intramolecular quadruplex was unwound.
猿猴病毒40（SV40）基因组是一个常用于研究真核生物复制的模型系统。大T抗原（T-ag）是一种病毒蛋白，负责在复制前解开SV40基因组并招募必要的宿主因子。除了双链解旋外，T-ag还具有G-四链体DNA解旋酶活性，其生理后果尚不清楚。然而，G-四链体DNA结构的形成可能参与基因组的维持和功能，而解开这些结构的解旋酶活性可能是高效复制所必需的。我们报告了首次使用表面等离子体共振（SPR）对SV40 T-ag解旋酶活性进行的实时研究。在ATP存在的情况下，观察到T-ag与固定化的单链DNA、叉状双链DNA以及人类端粒折叠四链体DNA序列结合。T-ag双链解旋酶活性的抑制可以实时观察到，并且分子内四链体被解开。
